colony pcr question

Steven Projan PROJANS at war.wyeth.com
Fri Jan 7 14:44:47 EST 2000


Not all gram positives are created equal and lysozyme can often be an inappropriate choice to crack open the cell wall.  For example Staphylococci are farily well imprevious to most muramidases but lysostaphin is quite effective, especially against the coagulase positive staph (e.g. S. aureus).  Another useful enzyme is mutanolysin which has some activity against staphylococci and is an important tool in breaking open the enterococci (sometimes used in combination with lysozyme).  Ogften doing a litle homework goes a long - check out volume 204 of Methods in Enzymology for some of this information as well publications describing work on the bacterium of interest.

Steve Projan


>>> Glen Tamura <gtamura at u.washington.edu> 01/06 6:41 PM >>>
Colony PCR on gram positive organisms can be quite different than from
gram negatives. In particular, gram positive cocci have a very thick cell
wall and often need to be digested with lysozyme in an appropriate buffer
before transferring a small quantity of this to the pcr reaction. 

Glen Tamura

On Thu, 6 Jan 2000, noodle wrote:

> I'm looking for any protocols for pcr'ing genomic dna from a colony of
> G+ bacteria
> 
> For example, do I need a boiling step before I begin the pcr? (etc.)
> 
> -Kelli
> 
> 
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